Nitrate: "the source makes the poison".

Interest in the role of dietary nitrate in human health and disease has grown exponentially in recent years. However, consensus is yet to be reached as to whether consuming nitrate from various food sources is beneficial or harmful to health. Global authorities continue to recommend an acceptable daily intake (ADI) of nitrate of 3.7 mg/kg-bw/day due to concerns over its carcinogenicity. This is despite evidence showing that nitrate consumption from vegetable sources, exceeding the ADI, is associated with decreased cancer prevalence and improvements in cardiovascular, oral, metabolic and neurocognitive health. This review examines the paradox between dietary nitrate and health and disease and highlights the key role of the dietary source and food matrix in moderating this interaction. We present mechanistic and epidemiological evidence to support the notion that consuming vegetable-derived nitrate promotes a beneficial increase in nitric oxide generation and limits toxic N-nitroso compound formation seen with high intakes of nitrate added during food processing or present in contaminated water. We demonstrate the need for a more pragmatic approach to nitrate-related nutritional research and guidelines. Ultimately, we provide an overview of our knowledge in this field to facilitate the various therapeutic applications of dietary nitrate, whilst maintaining population safety.

Characteristics of pulse oximetry and arterial blood gas in patients with fibrotic interstitial lung disease.

BACKGROUND: Fibrotic interstitial lung disease (ILD) is frequently associated with abnormal oxygenation; however, little is known about the accuracy of oxygen saturation by pulse oximetry (SpO2) compared with arterial blood gas (ABG) saturation (SaO2), the factors that influence the partial pressure of carbon dioxide (PaCO2) and the impact of PaCO2 on outcomes in patients with fibrotic ILD. STUDY DESIGN AND METHODS: Patients with fibrotic ILD enrolled in a large prospective registry with a room air ABG were included. Prespecified analyses included testing the correlation between SaO2 and SpO2, the difference between SaO2 and SpO2, the association of baseline characteristics with both the difference between SaO2 and SpO2 and the PaCO2, the association of baseline characteristics with acid-base category, and the association of PaCO2 and acid-base category with time to death or transplant. RESULTS: A total of 532 patients with fibrotic ILD were included. Mean resting SaO2 was 92±4% and SpO2 was 95±3%. Mean PaCO2 was 38±6 mmHg, with 135 patients having PaCO2 <35 mmHg and 62 having PaCO2 >45 mmHg. Correlation between SaO2 and SpO2 was mild to moderate (r=0.39), with SpO2 on average 3.0% higher than SaO2. No baseline characteristics were associated with the difference in SaO2 and SpO2. Variables associated with either elevated or abnormal (elevated or low) PaCO2 included higher smoking pack-years and lower baseline forced vital capacity (FVC). Lower baseline lung function was associated with an increased risk of chronic respiratory acidosis. PaCO2 and acid-base status were not associated with time to death or transplant. INTERPRETATION: SaO2 and SpO2 are weakly-to-moderately correlated in fibrotic ILD, with limited ability to accurately predict this difference. Abnormal PaCO2 was associated with baseline FVC but was not associated with outcomes.

Impact of Periodontitis on the Leakage of Oral Bacteria to the Gut.

Colorectal cancer (CRC) and periodontitis have recently been related due to the higher incidence of CRC in periodontal patients and the involvement of periodontal pathogens in carcinogenesis, suggesting that leakage from the oral cavity to the gut occurs. However, the magnitude of this pass-through in healthy individuals is controversial, and the effect that periodontitis could play in it is understudied. To evaluate the rate of bacterial leakage from the oral cavity to the gut, we analyzed the microbial composition of saliva, subgingival plaque, and fecal samples in healthy individuals without gastrointestinal disorders, including 20 periodontitis patients and 20 oral healthy controls, using PacBio full-length 16S rRNA gene sequencing. As expected, we observed a higher abundance of periodontal pathogens in the subgingival plaque and saliva of periodontal patients. In contrast, no significant differences were found between the fecal samples of both groups, implying that gut samples from periodontal patients were not enriched in periodontal pathogens. Fusobacterium nucleatum, a biomarker of CRC, was not found in the fecal samples of any participant. Our study does show a small leakage of some oral bacteria (mainly streptococci) to the gut, regardless of periodontal health status. Future studies should test whether other host factors and/or the preexistence of a gut disorder must be present in addition to periodontitis to promote the colonization of the gut by oral pathogens. The absence of periodontal pathogens in feces supports the idea that these bacteria could be used as biomarkers of intestinal disorders, including CRC.

Nasopharyngeal Swabs in Pediatric Patients With Thrombocytopenia and Anticoagulant Use.

BACKGROUND: Nasopharyngeal (NP) swabbing is a technique that is commonly used to test pediatric patients for viral infections with increased use during the coronavirus disease 2019 pandemic. Complications from NP swabbing are rare and seem to occur more frequently in patients at risk of bleeding. Little is known about institutional or individual practices and experiences with NP swab testing in pediatric patients with risk factors for bleeding. METHODS: We conducted a survey study of pediatric hematology/oncology (PHO) attending physicians to assess practices and experiences with NP swab testing in pediatric patients with thrombocytopenia and/or on anticoagulation. RESULTS: There were 130 total respondents (5.6%, n = 130/2327) from 6 countries. Relatively few respondents (n = 17/130, 13.1%) reported that their institution had a policy specifying a lower-level platelet cutoff for patients undergoing NP swabbing. The median platelet cutoff below which NP swabs are not performed according to existing policies is 30,000×10(9)/L (interquartile range: 20,000 to 40,000). The median cutoff based on the opinion of the respondents was 10,000 (interquartile range: 10,000 to 20,000). There were 24 episodes of epistaxis among PHO patients that were NP swabbed; many adverse events (56.5%, n = 13/23) were described as persistent, severe, and/or required intervention. Three reported cases of epistaxis with anticoagulation or antiplatelet therapy occurred in patients with concomitant thrombocytopenia. Only 1 respondent (n = 1/130, 0.7%) reported an institutional policy for limiting NP swabs in patients on anticoagulant therapy. NP (66.9%) and nares (33.1%) were the most common sources of coronavirus disease 2019 testing that were reported. CONCLUSION: A small percentage of institutions in this survey have a policy restricting NP swabs in PHO patients. The discrepancy between lower platelet cutoffs proposed by experts and institutional policy suggests that existing policies may be too conservative. Expert guidelines are needed on this topic. Other bleeding risk factors (eg, aspirin use and von Willebrand disease) should be considered in policies and guidelines.

An Effective Combination Therapy for the Treatment of Pediatric Monomorphic Post-transplant Lymphoproliferative Disorder With Plasmacytic Differentiation.

Post-transplant lymphoproliferative disorder (PTLD) is a complication of immunosuppressive therapy following solid organ or hematopoietic cell transplantation. Initial treatment typically includes a reduction of immunosuppression with or without rituximab. However, the optimal therapy for PTLD with plasmacytic differentiation is unclear. We present 3 cases of pediatric patients with plasmacytic PTLD. Two patients received a standard rituximab-based approach and relapsed, prompting additional chemotherapy. The third patient was treated with a novel regimen of bortezomib, dexamethasone, and daratumumab. This regimen was safe, well-tolerated, and resulted in a 2-year remission. Larger studies are needed to further explore this regimen.

The Effect of Surgeon Referral and a Radiation Oncologist Productivity-Based Metric on Radiation Therapy Receipt Among Elderly Women With Early Stage Breast Cancer: Analysis From a Tertiary Cancer Network.

Purpose: : Guidelines for early-stage breast cancer allow for radiation therapy (RT) omission after breast conserving surgery among older women, though high utilization of RT persists. This study explored surgeon referral and the effect of a productivity-based bonus metric for radiation oncologists in an academic institution with centralized quality assurance review. Methods and materials: : We evaluated patients ≥70 years of age treated with breast conserving surgery for estrogen receptor (ER)+ pT1N0 breast cancer at a single tertiary cancer network between 2015 and 2018. The primary outcomes were radiation oncology referral and RT receipt. Covariables included patient and physician characteristics and treatment decisions before versus after productivity metric implementation. Univariable generalized linear effects models explored associations between these outcomes and covariables. Results: : Of 703 patients included, 483 (69%) were referred to radiation oncology and 273 (39%) received RT (among those referred, 57% received RT). No difference in RT receipt pre- versus post-productivity metric implementation was observed (P = .57). RT receipt was associated with younger patient age (70-74 years; odds ratio [OR], 2.66; 95% confidence interval [CI], 1.54-4.57) and higher grade (grade 3; OR, 7.75; 95% CI, 3.33-18.07). Initial referral was associated with younger age (70-74; OR, 5.64; 95% CI, 3.37-0.45) and higher performance status (Karnofsky performance status ≥90; OR, 5.34; 95% CI, 2.63-10.83). Conclusions: : Nonreferral to radiation oncology accounted for half of RT omission but was based on age and Karnofsky performance status, in accordance with guidelines. Lack of radiation oncologist practice change in response to misaligned financial incentives is reassuring, potentially reflecting incentive design and/or centralized quality assurance review. Multi-institutional studies are needed to confirm these findings.

Tumor-associated microbiome features of metastatic colorectal cancer and clinical implications.

Background: Colon microbiome composition contributes to the pathogenesis of colorectal cancer (CRC) and prognosis. We analyzed 16S rRNA sequencing data from tumor samples of patients with metastatic CRC and determined the clinical implications. Materials and methods: We enrolled 133 patients with metastatic CRC at St. Vincent Hospital in Korea. The V3-V4 regions of the 16S rRNA gene from the tumor DNA were amplified, sequenced on an Illumina MiSeq, and analyzed using the DADA2 package. Results: After excluding samples that retained <5% of the total reads after merging, 120 samples were analyzed. The median age of patients was 63 years (range, 34-82 years), and 76 patients (63.3%) were male. The primary cancer sites were the right colon (27.5%), left colon (30.8%), and rectum (41.7%). All subjects received 5-fluouracil-based systemic chemotherapy. After removing genera with <1% of the total reads in each patient, 523 genera were identified. Rectal origin, high CEA level (≥10 ng/mL), and presence of lung metastasis showed higher richness. Survival analysis revealed that the presence of Prevotella (p = 0.052), Fusobacterium (p = 0.002), Selenomonas (p<0.001), Fretibacterium (p = 0.001), Porphyromonas (p = 0.007), Peptostreptococcus (p = 0.002), and Leptotrichia (p = 0.003) were associated with short overall survival (OS, <24 months), while the presence of Sphingomonas was associated with long OS (p = 0.070). From the multivariate analysis, the presence of Selenomonas (hazard ratio [HR], 6.35; 95% confidence interval [CI], 2.38-16.97; p<0.001) was associated with poor prognosis along with high CEA level. Conclusion: Tumor microbiome features may be useful prognostic biomarkers for metastatic CRC.

The tongue biofilm metatranscriptome identifies metabolic pathways associated with the presence or absence of halitosis.

Intra-oral halitosis usually results from the production of volatile sulfur compounds, such as methyl mercaptan and hydrogen sulfide, by the tongue microbiota. There are currently no reports on the microbial gene-expression profiles of the tongue microbiota in halitosis. In this study, we performed RNAseq of tongue coating samples from individuals with and without halitosis. The activity of Streptococcus (including S. parasanguinis), Veillonella (including V. dispar) and Rothia (including R. mucilaginosa) was associated with halitosis-free individuals while Prevotella (including P. shahi), Fusobacterium (including F. nucleatum) and Leptotrichia were associated with halitosis. Interestingly, the metatranscriptome of patients that only had halitosis levels of methyl mercaptan was similar to that of halitosis-free individuals. Finally, gene expression profiles showed a significant over-expression of genes involved in L-cysteine and L-homocysteine synthesis, as well as nitrate reduction genes, in halitosis-free individuals and an over-expression of genes responsible for cysteine degradation into hydrogen sulfide in halitosis patients.

Common human genetic variants of APOE impact murine COVID-19 mortality.

Clinical outcomes of severe acute respiratory syndrome 2 (SARS-CoV-2) infection are highly heterogeneous, ranging from asymptomatic infection to lethal coronavirus disease 2019 (COVID-19). The factors underlying this heterogeneity remain insufficiently understood. Genetic association studies have suggested that genetic variants contribute to the heterogeneity of COVID-19 outcomes, but the underlying potential causal mechanisms are insufficiently understood. Here we show that common variants of the apolipoprotein E (APOE) gene, homozygous in approximately 3% of the world's population1 and associated with Alzheimer's disease, atherosclerosis and anti-tumour immunity2-5, affect COVID-19 outcome in a mouse model that recapitulates increased susceptibility conferred by male sex and advanced age. Mice bearing the APOE2 or APOE4 variant exhibited rapid disease progression and poor survival outcomes relative to mice bearing the most prevalent APOE3 allele. APOE2 and APOE4 mice exhibited increased viral loads as well as suppressed adaptive immune responses early after infection. In vitro assays demonstrated increased infection in the presence of APOE2 and APOE4 relative to APOE3, indicating that differential outcomes are mediated by differential effects of APOE variants on both viral infection and antiviral immunity. Consistent with these in vivo findings in mice, our results also show that APOE genotype is associated with survival in patients infected with SARS-CoV-2 in the UK Biobank (candidate variant analysis, P = 2.6 × 10-7). Our findings suggest APOE genotype to partially explain the heterogeneity of COVID-19 outcomes and warrant prospective studies to assess APOE genotyping as a means of identifying patients at high risk for adverse outcomes.

Pediatric Germline Predisposition to Myeloid Neoplasms.

PURPOSE OF REVIEW: Advances in the understanding of germline predisposition to pediatric cancers, particularly myeloid neoplasms, have increased rapidly over the last 20 years. Here, we highlight the most up-to-date knowledge regarding known pathogenic germline variants that contribute to the development of myeloid neoplasms in children. RECENT FINDINGS: This discussion enumerates the most notable myeloid neoplasm-causing germline mutations. These mutations may be organized based on their molecular underpinnings-transcriptional control, splicing and signal transduction control, and a group of heterogeneous bone marrow failure syndromes. We review recent findings related to the biochemical mechanisms that predispose to malignant transformation in each condition. Key genetic discoveries such as novel mutations, degrees of penetrance, principles of the two-hit hypothesis, and co-occurrence of multiple mutations are shared. Clinical pearls, such as information regarding epidemiology, natural history, or prognosis, are also discussed. Germline mutations predisposing to pediatric myeloid neoplasms are frequent, but underrecognized. They hold major clinical implications regarding prognosis, treatment strategies, and screening for other malignancies. Further research is warranted to better characterize each of these conditions, as well as identify additional novel germline pathogenic variants of interest.

COVID-19-associated secondary hemophagocytic lymphohistiocytosis requiring hematopoietic cell transplant.

Coronavirus disease 2019 (COVID-19) infection causes a variety of extrapulmonary complications in pediatric patients. Multisystem inflammatory syndrome and hemophagocytic lymphohistiocytosis (HLH) are related to hypercytokinemia in COVID-19 patients. HLH is a disorder of exaggerated inflammation resulting in a cytokine storm and unrestricted hemophagocytosis. HLH can be primary (familial) or secondary (acquired). Secondary HLH (sHLH) can occur in patients with rheumatologic, oncologic, or infectious diseases. The link between COVID-19 and HLH has been reported in pediatric patients. Here we report a case of a pediatric patient who developed refractory sHLH secondary to COVID-19 infection and required a hematopoietic cell transplant for the cure.

Corona versus Dengue: Distinct Mechanisms for Inhibition of Polyprotein Processing by Antiviral Drugs.

Inhibitors interfering with processing of the viral polyprotein are used successfully for the control of extremely important viral pathogens, such as HIV and most recently SARS-CoV-2. This Viewpoint provides a mechanistic evaluation of a promising antiviral lead compound against dengue virus, JNJ-A07, 4-(3-((1-(4-chlorophenyl)-2-oxo-2-(6-(trifluoromethoxy)indolin-1-yl)ethyl)amino)-5-methoxyphenoxy)butanoic acid. The antiviral effect of JNJ-A07 appears, in our opinion, to be connected to an interference with the function of the viral protease. The analysis reveals for the first time that antiviral drugs target polyprotein processing not only by direct inhibition, but also by disturbing the native sequence of cleavage events. Implications on the development of broad-spectrum antivirals against flaviviruses are addressed.

RIP(K)ing away immunotherapy resistance.

Interferon signaling mediates resistance to immune checkpoint blockade therapy, but the underlying mechanisms are poorly understood. In this issue of Immunity, Cucolo et al. identify RIPK1 as an interferon-stimulated gene with potent effects on cell extrinsic and intrinsic immunotherapy resistance.

Prior COVID-19 infection may increase risk for developing endothelial dysfunction following hematopoietic cell transplantation.

Endothelial dysfunction underlies many of the major complications following hematopoietic cell transplantation (HCT), including transplant-associated thrombotic microangiopathy (TA-TMA), veno-occlusive disease/sinusoidal obstruction syndrome (VOD/SOS), and engraftment syndrome (ES). Emerging evidence similarly implicates endothelitis and microangiopathy in severe COVID-19-related multi-system organ dysfunction. Given the overlap in these two illness states, we hypothesize that prior COVID-19 infection may increase risk for HCT-related endotheliopathies. This retrospective, multicenter study included patients aged 0-25 years who underwent autologous or allogeneic HCT for any indication between January 1, 2020 and September 21, 2021, with close attention to those infected with COVID-19 in either the six months prior to transplant or twelve months following transplant. Incidences of TA-TMA, VOD/SOS, and ES were compared among patients with COVID-19 infection pre-HCT and post-HCT, as well as with historical controls who were never infected with SARS-CoV-2. Those who underwent HCT following COVID-19 infection displayed significantly increased rates of TA-TMA compared to those who were never infected. Additionally, our data suggests a similar trend for increased VOD/SOS and ES rates, although this did not reach statistical significance. Therefore, a history of COVID-19 infection prior to undergoing HCT may be a nonmodifiable risk factor for endothelial-related complications following HCT. Further studies are warranted to better clarify this relationship among larger cohorts and in the era of the Omicron SARS-CoV-2 variants.

Cryoprotection of humanin-like peptides in seminal plasma for ejaculated spermatozoa of crossbred bulls.

BACKGROUND: Cryopreservation process negatively affects spermatozoa functions. Humanin, a small polypeptide encoded in the mitochondrial genome, is well known for its role in cell survival. OBJECTIVE: To quantify the endogenous levels of humanin in seminal plasma of crossbred Frieswal bulls and to study its role in cryoprotection. The presence of humanin in bull spermatozoa was also investigated. MATERIALS AND METHODS: A total of 40 semen samples were separated into two groups based on the initial progressive motility (IPM): Good (IPM >70%) and Poor (IPM <50%) groups; and/or based on the post-thaw motility (PTM): Freezable (PTM>50%) and Non-freezable (PTM < 50%) groups. Humanin concentration in seminal plasma (SP-HN) was quantified using ELISA. RESULTS: SP-HN concentration ranged from undetectable to 67.6 pg/mL with a median level of 35.2 pg/mL. SP-HN level was significantly higher in the good quality semen group than in the poor quality semen group (p<0.001), and also significantly higher in the freezable group than in the non-freezable group (p<0.001). SP-HN level was positively correlated with initial progressive motility, post-thaw semen motility, viability, acrosome intactness and plasma membrane integrity, but negatively correlated the level of reactive oxygen species and malondialdehyde content. Immunochemical localization showed the presence of humanin in the proximal region of the middle piece of spermatozoa. CONCLUSION: Endogenous humanin level had significant correlation with semen quality and might protect sperm cells against freeze-induced oxidative stress. doi.org/10.54680/fr22510110712.

Spectrum of hematological malignancies, clonal evolution and outcomes in 144 Mayo Clinic patients with germline predisposition syndromes.

Germline predisposition syndromes (GPS) result from constitutional aberrations in tumor suppressive and homeostatic genes, increasing risk for neoplasia in affected kindred. In this study, we present clinical and genomic data on 144 Mayo Clinic patients with GPS; 59 evaluated prospectively using an algorithm-based diagnostic approach in the setting of a dedicated GPS/ inherited bone marrow failure syndrome (IBMFS) clinic. Seventy-two (50%) patients had IBMFS (telomere biology disorders-32,Fanconi anemia-18, Diamond Blackfan Anemia - 11, congenital neutropenia-5, Schwachman-Diamond Syndrome-5 and Bloom Syndrome-1), 27 (19%) had GPS with antecedent thrombocytopenia (RUNX1-FPD-15, ANKRD26-6, ETV6-2, GATA1-1, MPL-3), 28 (19%) had GPS without antecedent thrombocytopenia (GATA2 haploinsufficiency-16, DDX41-10, CBL-1 and CEBPA-1) and 17 (12%) had general cancer predisposition syndromes (ataxia telangiectasia-7, heterozygous ATM variants-3, CHEK2-2, TP53-2, CDK2NA-1, NF1-1 and Nijmegen Breakage Syndrome-1). Homozygous and heterozygous ATM pathogenic variants were exclusively associated with lymphoproliferative disorders (LPD), while DDX41 GPS was associated with LPD and myeloid neoplasms. The use of somatic NGS-testing identified clonal evolution in GPS patients, with ASXL1, RAS pathway genes, SRSF2 and TET2 being most frequently mutated. Fifty-two (91%) of 59 prospectively identified GPS patients had a change in their management approach, including additional GPS-related screening in 42 (71%), referral for allogenic HSCT workup and screening of related donors in 16 (27%), medication initiation and selection of specific conditioning regimens in 14 (24%), and genetic counseling with specific intent of fertility preservation and preconceptual counseling in 10 (17%) patients; highlighting the importance of dedicated GPS screening, detection and management programs for patients with hematological neoplasms.

[Mortality in patients with intracerebral haemorrhage associated with oral anticoagulation. Efficacy of a reversal protocol and clinical follow-up (ICH-OAC project)]. / Mortalidad en pacientes con hemorragia intracerebral asociada a anticoagulación oral. Eficacia de un protocolo de reversión y seguimiento clínico (proyecto HIC-ACO).

INTRODUCTION: Spontaneous intracerebral haemorrhage associated with oral anticoagulants (ICH-OAC) has a high mortality rate. The emergence of new anticoagulant drugs and reversal protocols increases interest in this entity. OBJECTIVES: The main objective is to determine the mortality rate in patients with ICH-OAC (early, in-hospital, global) in our health area and to analyse the main variables related to it. The secondary objective is to determine the efficacy of anticoagulation reversal therapies (ART) as reflected by radiological expansion of the haematoma and the functional prognosis. PATIENTS AND METHODS: A prospective observational study that introduced a protocol aimed at the management of patients with ICH-OAC. It included general measures and neuromonitoring, individualised administration of ART, cranial tomography and a six-month follow-up. Data on the drugs prescribed in the area during this period, mortality and functional prognosis were collected. A bivariate and logistic regression study was designed to investigate mortality-related variables. RESULTS: Forty-nine patients were included over three years; of these, 71.4% received ART. Mortality was 16.3% (first 24 hours), 53.1% (admission) and 61.2% (180 days). Lower survival was observed among patients with higher baseline scores on the National Institutes of Health Stroke Scale (NIHSS) (p < 0.0001), creatinine value (p = 0.02), International Normalised Index (p = 0.048), bleeding volume (p = 0.008), hydrocephalus (p = 0.015) and acenocoumarol intake (p = 0.030). Patients who did not receive ART had a greater rate of early mortality (p = 0.003). The only variable independently related to overall mortality was the baseline NIHSS score (odds ratio = 1.282; 95% confidence interval: 1.023-1.608; p = 0.031). CONCLUSIONS: ICH-OAC has a high mortality rate, related to the use of acenocoumarol and regardless of the initial clinical situation. A lower rate of early mortality was found among patients who received ART.

TITLE: Mortalidad en pacientes con hemorragia intracerebral asociada a anticoagulación oral. Eficacia de un protocolo de reversión y seguimiento clínico (proyecto HIC-ACO).Introducción. La hemorragia intracerebral espontánea asociada a anticoagulantes orales (HIC-ACO) presenta una elevada mortalidad. La aparición de nuevos fármacos anticoagulantes y protocolos de reversión aumenta el interés por esta entidad. Objetivos. El objetivo principal es determinar la tasa de mortalidad en pacientes con HIC-ACO (precoz, hospitalaria, global) en nuestra área sanitaria y analizar las principales variables relacionadas. El objetivo secundario es determinar la eficacia de las terapias de reversión de la anticoagulación (TRA), reflejada por la expansión radiológica del hematoma y el pronóstico funcional. Pacientes y métodos. Estudio prospectivo observacional que introdujo un protocolo dirigido al manejo de pacientes con HIC-ACO. Incluyó medidas generales y neuromonitorización, administración individualizada de TRA, tomografía craneal y seguimiento durante seis meses. Se recogieron los fármacos prescritos en el área durante este período, mortalidad y pronóstico funcional. Se diseñó un estudio bivariante y regresión logística para investigar variables relacionadas con la mortalidad. Resultados. Se incluyó a 49 pacientes durante tres años; de ellos, un 71,4% recibió TRA. La mortalidad fue del 16,3% (primeras 24 horas), el 53,1% (ingreso) y el 61,2% (180 días). Se observó una menor supervivencia entre pacientes con puntuaciones basales mayores en la National Institutes of Healt Stroke Scale (NIHSS) (p lower than 0,0001), valor de creatinina (p = 0,02), índice internacional normalizado (p = 0,048), volumen hemorrágico (p = 0,008), hidrocefalia (p = 0,015) y toma de acenocumarol (p = 0,030). Los pacientes que no recibieron TRA tuvieron una mayor mortalidad precoz (p = 0,003). La única variable relacionada con la mortalidad global de forma independiente fue la puntuación en la NIHSS basal (odds ratio = 1,282; intervalo de confianza al 95%: 1,023-1,608; p = 0,031). Conclusiones. La HIC-ACO presenta una elevada mortalidad, relacionada con la toma de acenocumarol y de forma independiente con la situación clínica inicial. Se comprobó una menor tasa de mortalidad precoz entre pacientes que recibieron TRA.

Efficiency Improvements and Discovery of New Substrates for a SARS-CoV-2 Main Protease FRET Assay.

The COVID-19 pandemic, caused by the SARS-CoV-2 virus, has a huge impact on the world. Although several vaccines have recently reached the market, the development of specific antiviral drugs against SARS-CoV-2 is an important additional strategy in fighting the pandemic. One of the most promising pharmacological targets is the viral main protease (Mpro). Here, we present an optimized biochemical assay procedure for SARS-CoV-2 Mpro. We have comprehensively investigated the influence of different buffer components and conditions on the assay performance and characterized Förster resonance energy transfer (FRET) substrates with a preference for 2-Abz/Tyr(3-NO2) FRET pairs. The substrates 2-AbzSAVLQSGTyr(3-NO2)R-OH, a truncated version of the established DABCYL/EDANS FRET substrate, and 2-AbzVVTLQSGTyr(3-NO2)R-OH are promising candidates for screening and inhibitor characterization. In the latter substrate, the incorporation of Val at position P5 improved the catalytic efficiency. Based on the obtained results, we present here a reproducible, reliable assay protocol using highly affordable buffer components.